PROJECT SUMMARY - PROJECT 2 The central goal of Project 2 is to understand the various roles of alternative splicing in cancer, and to exploit cancer-specific features of this process to develop targeted-therapeutic approaches. Cancer cells display extensive qualitative and quantitative dysregulation of splicing, and a subset of the numerous isoforms that are inappropriately expressed contribute to tumorigenesis or altered cell metabolism. The mechanisms and pathways through which the splicing-factor oncoproteins SRSF2 and SRSF1 transform cells will continue to be investigated. Cell-culture models, as well as orthotopic and genetic mouse models will be used to study tumorigenesis promoted by these splicing factors upon mutation or overexpression in different cancer contexts, with an emphasis on recurrent mutations in myeloid dysplasias. High-throughput RNA-sequencing and computational analysis will be employed to identify and compare the splicing targets of these SR proteins in different cancer contexts, and selected targets will be characterized and manipulated to evaluate their contributions to tumorigenesis and potential as therapeutic targets or biomarkers. One key event, alternative splicing of pyruvate kinase pre-mRNA, which controls the distinctive glycolytic metabolism of cancer cells, will be thoroughly investigated as a potential therapeutic target, by specifically manipulating this process in vivo, using antisense technology and mouse models of glioma and hepatocellular carcinoma.